Combined effects of chlorpromazine and high fat diet on lipid levels and enzyme activities in bile, liver and plasma of Wistar rats

The effects of high fat diet and injection of chlorpromazine on bile lipid secretion were studied in the rats fed a control diet (C), a saturated fat, high cholesterol diet (S) and a polyunsaturated fat, high cholesterol diet (PU). As compared to controls, injection of chlorpromazine in the S and PU diet groups caused no appreciable change in the level of bile salts and bile phospholipids. Chlorpromazine did however enhance bile cholesterol, especially in the PU group, and lower secretion of lysosomal enzyme (beta-glucuronidase) into bile. Impairment of lysosomal enzyme secretion but not of bile lipid secretion suggests that the lysosomal activity is not directly involved in the bile secretion mechanism. These data point up the risks of using chlorpromazine therapy in association with a diet high in fat and cholesterol.     

Proteoglycan synthesis by normal and neoplastic human transitional epithelial cells

Metabolically 35S-labeled proteoglycans were isolated from cell-associated matrices and media of confluent cultures of human normal transitional epithelial cells and HCV-29T transitional carcinoma cells. On Sepharose CL-4B columns, the cell-associated proteoglycans synthesized from both cell types separated into three identical size classes, termed CI, CII, and CIII. Normal epithelial cell C-fractions eluted in a 22:34:45 proportion and contained 64%, 64%, and 72% heparan sulfate, whereas corresponding HCV-29T fractions eluted in a 29:11:60 proportion, and contained 91%, 77%, and 70% heparan sulfate, respectively. Medium proteoglycans from normal cells separated into two size classes in a proportion of 6:94 and were composed of 35% and 50% heparan sulfate. HCV-29T medium contained only one size class of proteoglycans consisting of 23% heparan sulfate. The remaining percentages were accounted for by chondroitin/dermatan sulfate. On isopycnic CsCl gradients, proteoglycan fractions from normal cells had buoyant densities that were higher than the corresponding fractions from HCV-29T cells. DEAE-Sephacel chromatography showed that cell and medium associated heparan sulfate from HCV-29T cells was consistently of lower charge density (undersulfated) than that from normal epithelial cells. In contrast, the chondroitin/dermatan sulfate of HCV-29T was of a charge density similar to that of normal cells. These as well as other structural and compositional differences in the proteoglycan may account, at least in part, for the altered behavioral traits of highly invasive carcinoma cells.     

Antigen-presenting cell-T cell interaction in the chicken is MHC class II antigen restricted

The involvement of the MHC in the recognition of Ag by avian T lymphocytes was analyzed. PBL from chickens primed with keyhole limpet hemocyanin in vivo were induced to synthesize DNA in an in vitro response to specific Ag. Responding cells were T cells as judged by immunofluorescence staining. In vivo Ag-primed PBL were stimulated in vitro with specific Ag and further propagated in the presence of IL-2. Subsequent Ag-specific T cell proliferation required the presence of Ag-pulsed peripheral blood adherent cells (APC). T cell responses were restricted by the MHC of the APC; Ag presented by allogeneic APC did not support T cell proliferation. By using MHC-recombinant chicken lines, the gene products controlled by MHC class II loci were shown to restrict the T cell-APC interaction. This conclusion was substantiated by the inhibition of the Ag-specific T cell response by a mAb against chicken MHC class II gene products but not by a mAb against chicken MHC class I gene products.     

Induction of reversible changes in cell-surface glycoconjugates and lung colonization potential by 13-cis retinoic acid

Murine squamous carcinoma cells (KLN205) grown in a medium supplemented with the retinoid, 13-cis retinoic acid (RA), had dose-dependent, selective increases in the expression of certain lectin receptors, which correlated with a dramatic decrease in the ability to form pulmonary colonies (P ?.0003) (Couch MJ, Pauli BU, Weinstein RS, Coon JS: JNCI, 78:971 ?977, 1987). These findings suggest a possible relationship between the RA-induced glycoconjugate alterations and the decreased experimental metastatic behavior. We further define the mechanism of RA's action. The finding that RA treatment (5 × 10^?6 M, 5 × 10^?7 M) did not perturb the cell cycle of KLN205 cells provides further proof that the decreased metastatic behavior is not attributable to any inhibition in the rate of growth or to alterations in the cell cycle. Furthermore, since stable subpopulations with variable lectin binding could not be detected, the mechanism of RA's action does not appear to be due to selection of variant tumor-cell subpopulations. Finally, in a scries of experiments designed to determine the reversibility of the RA treatment, the RA-induced decrease in metastatic behavior reverted back to a more metastatic state in the same time frame (3 days) as the reversion of the RA-induced changes in cell-surface glycoconjugate expression. This reversion provides further evidence for a close relationship between the RA-induced modulation of tumor cell-surface glycoconjugate expression and the decreased metastatic behavior; it suggests that transient, reversible modulation of the tumor cell surface may play a role in determining metastatic behavior.     

Characterization of murine monoclonal antibodies directed against the core proteins of human immunodeficiency virus types 1 and 2.

Monoclonal antibodies (MAbs) raised against the core proteins of human immunodeficiency virus type 1 (HIV-1; laboratory strain HTLV-IIIB) and HIV-2 (strain ROD) were investigated in a variety of tests, e.g., enzyme-linked immunosorbent assay (ELISA), immunostaining of Western immunoblots, immunofluorescence, immunoprecipitation, and alkaline phosphatase anti-alkaline phosphatase assay. The MAbs were grouped according to their cross-reactions. Seven HIV-1-specific MAbs reacted exclusively with HIV-1, and five showed cross-reactivity with HIV-2 and simian immunodeficiency virus of macaques in ELISA. Four of the 15 MAbs against HIV-2 reacted only with the HIV-2 protein p26. Six showed cross-reactivity with HIV-1, and five showed a broad reaction with all three viruses. Overlapping 30-amino-acid-long peptides derived from the p24 protein sequence of HIV-1 were used in an epitope-mapping system. Three different immunogenic regions (A, B, and C) could be defined. Specific regions where anti-HIV-1 and -HIV-2 MAbs cross-reacted were mapped with shorter oligopeptides.     

Characterization of HSP27 and three immunologically related polypeptides during Drosophila development

The low-molecular-weight heat-shock protein HSP27 is made in the absence of heat shock during Drosophila melanogaster development. An analysis of the accumulation of HSP27 during specific stages of development is presented using an antiserum recognizing this protein. Whereas HSP27 is abundant during embryogenesis, the level of this protein begins to decrease in the 20-h old embryo and is no longer detectable in second instar larvae. A high level of HSP27 is again observed in third instar larvae and reaches a maximal level in late pupae. While still abundant in young adult flies of both sexes, a greater amount of HSP27 is found in females with the protein being highly concentrated within the ovaries. Following lysis of whole pupae, about 60% of HSP27 is found in the soluble lysate fraction in a form which sediments between 5 and 20 S. Anti-HSP27 serum also recognizes three other developmentally regulated polypeptides with apparent MW of 33, 85 and 120 kDa. The 33 kDa protein accumulates in pupae while those of 85 and 120 kDa are more abundant in third instar larvae. Unlike HSP27, these proteins are not detected in embryos or ovaries. Immunoblot analysis of V8 proteolytic fragments suggests that HSP 27 and 33 kDa are related polypeptides. Exposure of the developing insect to heat-shock treatment results in increased level of HSP27. In larvae, a small amount of the 33 kDa protein accumulates following heat shock, while in pupae and adult flies a decrease in the concentration of this protein is observed after heat shock. Finally, different cellular localizations and distributions within the pupal body have been found for these developmentally regulated polypeptides.     

The comparative influence of substituted phenols (especially chlorophenols) on yeast cells assayed by electro-rotation and other methods

The toxicity of 31 phenols was studied by electro-rotation of yeast cells. Control yeast cells show both anti-field and co-field rotation, depending upon the field frequency applied. After treatment with supra-threshold amounts of phenols the anti-field rotation is weakened or abolished and a stronger co-field rotation can be seen. The proportion of cells showing the co-field rotation was found to be a sensitive measure of toxicity. Doses of 2.2 mumol/l of pentachlorophenol, or of 0.3 mumol/l of pentabromophenol were detectable after 3 h incubation at pH 4.0. At a given pH, the toxicity of the chlorophenols correlated extremely well with their octanol:water partition coefficients (Pow). The complete set of phenols showed fair overall correlation with Pow, but less good correlation with their acidity constants (pKa). In particular the toxicity of a given phenol was less than predicted from its pKa if the incubation pH was higher than the pKa. Biochemical assays on 23 of the phenols showed that the rotational sensitivity runs closely parallel to the sensitivities of cell growth rate and of the plasmamembrane ATPase, but less closely to the inhibition of purine incorporation. It appears that the electro-rotation method provides a useful and rapid test for the presence of organic ecotoxins. The test enables us to distinguish differences between single cells, and is comparable in sensitivity to biochemical tests that use vesicles or homogenates derived from a cell population.     

Hydrolysis of phospholipid monolayers by human spermatozoa. Inhibition by male contraceptive gossypol

Monomolecular films of phospholipid were used to study the interaction of intact human spermatozoa with model membranes. Exclusively with negatively charged phosphatidylglycerol monolayers rapid penetration of spermatozoa into the monolayer with subsequent hydrolysis of the lipid was triggered by the addition of 5 mM calcium into the medium. The results suggest the localization of a calcium-dependent phospholipase A2 at the outer acrosomal or plasma membrane of human spermatozoa with its active site exposed to the external environment. Preincubation of the cells with 100 microM gossypol completely abolished the ability of human spermatozoa to hydrolyze or penetrate monolayers of phosphatidylglycerol. The inhibition of the phospholipase activity by gossypol may contribute to the unknown contraceptive mechanisms of this non-steroidal male antifertility agent.     

Acute effects of filipin on the plasmic, hepatic, and biliary cholesterol of the rat

Twenty-one male Wistar rats, 13 weeks old, were fed ad libitum hyperlipidic diets (28% fats) loaded with cholesterol (1.2%) for 5 weeks. One group of 11 rats was fed saturated fats (diet group "S") and another group of 10 rats was fed polyunsaturated fats (diet group "PU"). On the day they were sacrificed 10 of the rats were injected intravenously with 1 mg of filipin. Contrary to the rats in diet group "PU," the rats in diet group "S" treated with filipin presented certain characteristics that were not found in the nontreated group: They provided evidence of biliary cholestasis accompanied by a decline in the level of secretion of bile salts and phospholipids into bile. The concentrations of both free and esterified cholesterol in plasma fell and the amount of (esterified) hepatic cholesterol rose, although there was no change due to the filipin in the amounts of hepatic phospholipids. Explanatory hypotheses for these phenomena were considered, first, at the site of plasma membranes where filipin binds selectively to the cholesterol in the membrane, causing a disruption which probably disturbs the absorbance of circulating lipoproteins, especially that of hepatocyte cells, particularly in diet group "PU." Second, the effects of filipin on subcellular membranes seem to disturb the secretion of lipids and lipoproteins into bile and plasma, especially in diet group "S." Last, at the intracellular level, filipin appears to have a blocking effect on the organelles involved in biliary lipid secretion. The activity of certain enzymes such as cholesterol esterase may also be blocked, particularly in diet group "S," which would explain the accumulation of esterified cholesterol in liver.     

Increased endogenous retroviral gene expression is a consequence of lymphocyte activation

Plasma cells of line 151(5) chickens have been shown to express elevated levels of endogenous retroviral envelope glycoprotein (VEG), measured relative to levels expressed by both immature B cells and resting peripheral B lymphocytes. In this study we analyzed the relationship between peripheral blood lymphocyte (PBL) maturation and the level of VEG expression. A culture system was developed that would support maturation of pokeweed mitogen-activated peripheral B lymphocytes. As analyzed by cytofluorometry, both Ig+ and Ig- lymphoblasts present in the pokeweed mitogen-stimulated cultures expressed detectable levels of VEG in contrast to bursacytes and PBL. Similarly, Ig- blasts, which were present in concanavalin A-stimulated cultures of PBL and presumed to represent activated T cells, were also positive for the expression of VEG. Immature T cells, i.e., thymocytes, although negative by immunofluorescence analysis, expressed VEG at levels that were detectable by radioimmunochemical techniques. These results indicate that T cells as well as B cells constitutively express VEG, and that mitogenic activation of the resting lymphocyte induces an increase in VEG expression.